| Picea wilsonii is an evergreen coniferous tall tree of Pinaceae.It is a unique tree species in China and has the characteristics of drought resistance and strong adaptability.At present,with the continuous deterioration of the environment and the interference of human factors,the population and distribution range of the Picea wilsonii drastically decrease,and through the research on the resistance mechanism of indigo mysica,especially related stress resistance genes,the population of barley can be recovered.And distribution,laying a theoretical foundation for retaining high-quality stress-resistant genes in trees.Salt stress is one of the important abiotic stress factors affecting agroforestry production in the world,and the use of plant biology related technologies such as genetic engineering breeding and molecular assisted breeding to improve the tolerance of forest trees and crops to salt stress is high-quality and high-quality method.In this paper,four genes related to salt stress response were screened from the transcriptome library of the indica striolata using bioinformatics and qRT-PCR experiments.In order to further verify the related functions of the target gene,potato of ’Atlantic’cultivar was selected as the genetic transformation material.The screening of the potato induction medium and Agrobacterium tumefaciens genetic transformation conditions was performed to establish a suitable gene function verification system.The results are as follows:(1)Four genes that responded to salt stress,namely PwWRKY1,PwTLP1,PwSCP1,and PwLRR1,were screened from the transcriptome library of the indigo indica and analyzed by bioinformatics.The PwWRKY1 and PwLRR1 genes had the highest expression levels in roots,while PwTLP1 had the highest expression levels in leaves and roots.These three species also had the lowest expression levels in pollen.PwSCP1,unlike the other three,had the highest expression in stems and the lowest in roots.(2)Analysis of gene expression patterns under salt,dry,cold,and ABA stress conditions indicated that the three target genes PwWRKY1,PwSCP1,and PwLRR1 were all significantly involved in the stress process of the above 4 stress conditions;whereas the PwTLP1 gene was only There was a significant up-regulated expression trend for salt and drought stress,but no significant response to cold and ABA conditions.(3)Potatoes of the ’Atlantician’ variety were selected as experimental materials in this experiment for potato-induced tuber experiments.The results showed that Formula 3(80g/L Sucrose+2mg/L 6-BA),although it induces tuberculosis The rate is only ranked 3rd,which is 92%,but its large potato rate and large number of potatoes far exceeds other combinations,reaching 76.I%.Therefore,given the need for subsequent genetic transformation experiments,it is considered that the No.3 tuber formula is most suitable for Induction of the ’Atlantician’ variety in vitro.(4)In this experiment,the genetic transformation conditions of the most suitable ’Atlantician’potato chips were screened,and the results showed that the transformation conditions of the fifth group(the infection 8min,the OD value of 0.8,Hyg concentration of lOmg/L)has the highest the conversion rate which significantly higher than other combinations,reaching 45.167%,and its browning rate was also significantly lower than other combinations,only 35.767%.Therefore,this combination is considered to be the most suitable combination of transformation conditions for genetic transformation of ’Atlantic’ potato chips.The influence degree of each factor on the conversion efficiency is:Hyg concentration>Dyeing time>OD value.(5)In this experiment,four genes related to salt stress response were cloned from PwWRKY1,PwTLP1,PwSCP1,and PwLRR1.Potato chips were transformed by Agrobacterium inoculation,and 4 PwWRKYI and 2 PwLRR1 genes of transgenic positive vaccines were obtained.The seedlings provide the basis for further verification of their gene functions. |
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